Streptococcus mutans is the primary causative agent of dental caries, which remains the most common chronic disease among children and is untreated in up to 30% of adults over 35 yrs-old. When transiently introduced into the bloodstream following daily dental hygienic practices such as tooth brushing and flossing, this bacterium can also cause potentially lethal endocarditis infections in atrisk patients. S. mutans virulence is intimately related to its ability to form biofilm, but the molecular mechanisms facilitating its initial attachment to the tooth surface, as well as biofilm maturation and dispersal, are not well understood. In this respect, cell death and lysis are aspects of bacterial physiology that have been implicated in S. mutans biofilm formation. Although well-studied in other microorganisms, the molecular mechanisms that regulate cell death and autolysis in S. mutans have not been clearly defined. To this end, a developmental research project is proposed that initiates study of the cid and lrg genes in S. mutans, as these genes have been shown to be important regulators of cell death, autolysis and biofilm development in the pathogen Staphylococcus aureus. The specific aims of this application are (1) to ascertain the role of the cid and lrg genes in regulating S. mutans cell death and lysis during biofilm development, and (2) to examine the genetic and metabolic factors that control expression of the cid and lrg genes in S. mutans. Allele replacement mutations will be created in the cid, lrg, and lytS genes in both laboratory and clinical S. mutans strains. The effect of these mutations on cell death and lysis will be measured by a variety of phenotypic assays. Confocal microscopy will be used to assess their effect on biofilm adherence and development in flow cell and static models, as well as to monitor fluorescence of cid and lrg GFP promoter fusions. The role of metabolic signals and the lytS gene on cid and lrg expression will also be assessed. It is anticipated that this research will delineate a critical aspect of S. mutans physiology and pathogenesis that will help improve strategies of prevention and treatment of dental caries, as well as infective endocarditis. This research is also an important first step in achieving the long-term goal of defining the mechanisms that control cell death and autolysis in the oral streptococci, and how these processes contribute to their ability to grow and survive in dental plaque as well as their ability to colonize heart valves during endocarditis. PUBLIC HEALTH RELEVANCE: Despite wide-spread implementation of preventative measures such as community water fluoridation and school-based dental sealant programs, dental caries remains the most common chronic disease among children, and is also prevalent among adults. Streptococcus mutans, the bacteria that causes dental caries, can also cause infective endocarditis in at-risk patients. Therefore, research that contributes to our understanding of how this pathogen is able to colonize and grow in dental plaque will facilitate the development of novel preventative and/or treatment strategies for both of these diseases.